Anti-Blue Antibody 100ul - dilute 1:2000 (100ul - dilute 1:2000)

With Anti-Blue antibody the time of manually marking your protein marker bands is forever gone. No more guessing how accurately the marker bands were charted! Anti-Blue visualizes your markers and your target proteins together on the same blot, in the same exposure.

The indicated amounts of the respective protein markers were separated by SDS-PAGE and transferred to nitrocellulose membrane. The membrane was blocked with 3% skim milk in PBS-T for 1 h at room temperature and incubated with Anti-Blue antibody diluted 1:2000 and secondary anti-mouse HRP in 0.5% skim milk in PBS-T for 1 h at room temperature. The membrane was washed 3x with PBS-T and processed for ECL detection.

The Anti-Blue antibody has been extensively tested with whole cell lysates of different species, confirming the extremely high specificity towards blue-stained proteins. No cross-reactivity with whole cell proteins has been detected

M.....Molecular Weight Marker

1......Mouse Embryo Fibroblasts

2......U2OS

3......Rat1

4......BHK21

5......CV1

6......S. cerevisiae (BY4741)

7......E. Coli (Rosetta(DE3)pLysS)

8......Chicken follicle

Anti-Blue antibody can be used at the primary antibody probing stage without interfering with the identification of the target protein. Anti-Blue is diluted 1:2000 into the primary antibody solution and applied to the membrane followed by detection with an anti-mouse secondary antibody. Alternatively Anti-Blue antibody can be used directly with secondary anti-mouse antibodies (diluted 1:2000)

1... U2OS 4HA-B56β

2... NIH3T3 myc-Cα/HA-B55α

3... NIH3T3 myc-Aα

4... MEF PME1-/-

M... Molecular Weight Marker (1μl)

HRP-Labelled Anti-Blue

Where the primary antibody prevents the use of anti-mouse secondary antibodies, HRP-labelled Anti-Blue can be used. This antibody can be diluted 1:2000 with a secondary antibody of choice.

Comparison of label free Anti-Blue vs HRP-Labelled Anti-Blue

The indicated volumes of 2-Color SDS Marker were separated by SDS-Page and transfered to nitrocellulose membrane. The membranes were blocked with 3% dry milk in PBS-Tween for 1h at room temperature and incubated with either

• •  The label-free Anti-Blue antibody diluted 1:2000 + anti-mouse HRP labelled secondary antibody or;

• •  HRP-labelled Anti-Blue antibody diluted 1:2000 in 0.5% dry milk in PBS-Tween for 2h at RT.

Detection was performed with ECL Western Blotting Detection Reagents.

Freeze Thaw

The Anti-Blue antibody shows excellent resistance to repeated freeze/thaw cycles which enables repeated and frequent use without the need to aliquot smaller samples. Aliquots of Anti-Blue, were repeatedly frozen at -20°C and thawed on ice. Nitrocellulose membranes containing the indicated volumes of pre-stained marker were blocked with 3% dry milk in PBS-Tween for 1h at room temperature and incubated with the Anti-Blue diluted 1:2000, together with anti-mouse HRP labelled secondary antibody in 0.5% dry milk in PBS-Tween for 2h at room temperature. Detection was performed with ECL Western Blotting Detection Reagents. No decrease in sensitivity was detected after 5 freeze thaw cycles.